Transmission electron microscopy is a versatile tool widely used across a range of research areas from biology to physio-chemical subjects. The transmission electron microscope is one of the most effective imaging devices available and can be used to study not only the surface morphology but also the internal structure and defects in diverse materials including metals, semiconductors, proteins and polymers.
A schematic diagram illustrating the workings of a transmission electron microscope (TEM) is shown in FIG. 1. In essence, a TEM consists of an electron gun 1 which produces a high-energy beam of electrons, and a series of electromagnetic lenses 2 which control and collimate the beam along the optic axis A-A onto a specimen under investigation. A thin sample 4 (˜100 nm thick) is used so that electrons may be transmitted through the material. The electrons are scattered by the sample and form a diffraction pattern characteristic of that material. A further series of lenses 5 can be used to form an image from the electron beams that had been scattered as they passed through the sample. The image or diffraction pattern may be viewed on a screen 6 coated with electron-fluorescent material, through a viewing window 7, or converted to a picture shown on a monitor via camera chamber 8. Alternatively, techniques for quantitative measurement of the electron intensity across the image may be employed. The whole TEM cavity is evacuated to a pressure of less than 10−2 Pa, to prevent the beam of electrons being disrupted by gas particles.
In order to obtain the high resolution that is often required in a TEM, the sample is generally positioned within the electromagnetic lenses or “pole pieces” 2c and 5a. The gap inside a high-resolution pole piece is small and this places a severe restriction on its maximum size: typically the sample 4 is a disc, 3 mm in diameter and with a maximum height of approximately 1 mm (FIG. 2). In order that electrons may be transmitted through the sample, specialist sample preparation techniques are used to thin the material to less than 100 nanometres. In most techniques, material is removed from the center of the sample disk using electro-polishing methods or ion thinning until a hole 10 is made in the center of the disk. A thin, electron-transparent area 11 surrounds the hole.
The prepared sample 4 is placed in a specimen holder 3 which extends through the wall of the TEM to hold the sample in the required location. The end of the specimen holder in which the sample is held is the specimen tip. This is generally an integral part of the specimen holder 3. A variety of specimen tips are available, examples of which are shown in FIGS. 3a, 3b and 3c. The sample is generally held in a circular recess 12 in the specimen tip, provided with a central aperture 13 intended to coincide with the hole 10 and surrounding electron-transparent region 11 of the sample 4. Often the tip is equipped with a thin mesh or grid which provides the sample with further support. Finally, the sample is held in place by a clip ring or similar mechanical fastening means. Available holders include heating and cooling stages, electrical stages which measure voltage and current in the specimen and straining stages. Examples of such stages are described in U.S. Pat. No. 5,225,683 which discloses various types of specimen tip with conventional clamping and tilting means which can be interchangeably mounted to a specimen holder.
Certain specialised techniques exist for mounting particular types of sample. For example, GB-A-2121208 describes a technique for freeze-drying and mounting cryosections. A pressure element is used to locate the specimen and, due to the applied pressure, the specimen becomes affixed to the mount during drying.
By combining a series of TEM images of a sample, it is possible to create a three-dimensional model of the specimen. This is termed “electron tomography” and is an important research tool since many complex specimens cannot be adequately described by a two-dimensional projection alone. For example, electron tomography is often used to image small single particles, such as catalysts or viruses.
In order to obtain the series of images required to form a 3-D model, the sample must be tilted through as large an angle as possible. Conventional specimen holders as described above, including those of U.S. Pat. No. 5,225,683, are bulky and, in the confined space within a high-resolution pole piece, do not generally allow the sample to be tilted by more than +/−40°. It would be advantageous for the holder size to be reduced so that the tilt range could be increased.
Furthermore, when tilting the sample it is important that the area of interest on the sample does not move out of focus or out of the viewing field. This is achieved by positioning the sample such that the point of interest is at the same height as the tilt axis. This is termed the “eucentric height”. Conventionally standard holders are designed such that a standard size and thickness of sample will be approximately at the eucentric height when placed in the TEM. The height of the specimen holder relative to the TEM may be adjusted for focus in a goniometer which provides movement of the holder in 3 directions (x, y, z) with respect to the electron optics of the TEM. This is disclosed, for example, in JP2001-068047. However, due to the variation in the thickness of the sample, this technique does not guarantee that the sample is at the eucentric point and it would be advantageous if the z-axis height of the sample could be fine-tuned by the user.
U.S. Pat. No. 3,778,621 discloses a specimen tilting device for an electron microscope. The device provides for tilting of a mounted specimen about the X and Y axis, and lateral motion in the X axis (parallel to the axis of the device). The specimen stage may also be pivoted so as to move the specimen along an arcuate path in the XY or YZ planes. However the device is of a complex and delicate construction and does not provide the ability to fine-tune the Z-axis position of the specimen without moving the specimen in the XY plane also,